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1.
J Biol Chem ; 300(1): 105564, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38103644

RESUMO

The polysialyltransferases ST8SIA2 and ST8SIA4 and their product, polysialic acid (polySia), are known to be related to cancers and mental disorders. ST8SIA2 and ST8SIA4 have conserved amino acid (AA) sequence motifs essential for the synthesis of the polySia structures on the neural cell adhesion molecule. To search for a new motif in the polysialyltransferases, we adopted the in silico Individual Meta Random Forest program that can predict disease-related AA substitutions. The Individual Meta Random Forest program predicted a new eight-amino-acids sequence motif consisting of highly pathogenic AA residues, thus designated as the pathogenic (P) motif. A series of alanine point mutation experiments in the pathogenic motif (P motif) showed that most P motif mutants lost the polysialylation activity without changing the proper enzyme expression levels or localization in the Golgi. In addition, we evaluated the enzyme stability of the P motif mutants using newly established calculations of mutation energy, demonstrating that the subtle change of the conformational energy regulates the activity. In the AlphaFold2 model, we found that the P motif was a buried ß-strand underneath the known surface motifs unique to ST8SIA2 and ST8SIA4. Taken together, the P motif is a novel buried ß-strand that regulates the full activity of polysialyltransferases from the inside of the molecule.


Assuntos
Mutação , Sialiltransferases , Humanos , Motivos de Aminoácidos/genética , Substituição de Aminoácidos , Simulação por Computador , Complexo de Golgi/enzimologia , Complexo de Golgi/metabolismo , Moléculas de Adesão de Célula Nervosa/química , Moléculas de Adesão de Célula Nervosa/metabolismo , Mutação Puntual , Conformação Proteica em Folha beta , Transporte Proteico , Algoritmo Florestas Aleatórias , Ácidos Siálicos/metabolismo , Sialiltransferases/química , Sialiltransferases/genética , Sialiltransferases/metabolismo
2.
Dev Comp Immunol ; 159: 105224, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-38969190

RESUMO

Stimulator of interferon genes (STING) mediates innate immune response upon binding to cyclic GMP-AMP (cGAMP). It recruits tank-binding kinase 1 (TBK1) and transcription factor interferon regulatory factor 3 (IRF3) through its C-terminal tail and facilitates TBK1-dependent phosphorylation of IRF3 via forming STING polymers in mammalian cells. However, the mechanism behind STING-mediated activation of NF-κB transcription factor, Relish, in insect cells is unknown. Our study revealed that insect STING formed oligomers and the cryptic RIP homotypic interaction motif (cRHIM) was required for its oligomerization and its anti-viral functions. Cells expressing cRHIM-deficient mutants exhibited lower levels of anti-viral molecules, higher viral load after viral infection and weak activation of Relish. Moreover, we observed that under cGAMP stimulation, insect STING interacted with IMD, and deletion of the cRHIM motif on either protein prevented this interaction. Finally, we demonstrated that cGAMP enhanced the amyloid-like property of insect STING aggregates by ThT staining. In summary, our research showed that insect STING employed a homotypic motif to form intermolecular interactions that are essential for its antiviral signaling.


Assuntos
Imunidade Inata , Fator Regulador 3 de Interferon , Proteínas de Membrana , Transdução de Sinais , Animais , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Transdução de Sinais/imunologia , Fator Regulador 3 de Interferon/metabolismo , Fator Regulador 3 de Interferon/genética , Nucleotídeos Cíclicos/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas de Drosophila/metabolismo , Proteínas de Drosophila/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Motivos de Aminoácidos/genética , Humanos , Linhagem Celular , Ligação Proteica , Fosforilação , Multimerização Proteica , Drosophila melanogaster/imunologia , Drosophila melanogaster/virologia
3.
Braz. j. med. biol. res ; 47(5): 369-375, 02/05/2014. tab, graf
Artigo em Inglês | LILACS | ID: lil-709431

RESUMO

To investigate signal regulation models of gastric cancer, databases and literature were used to construct the signaling network in humans. Topological characteristics of the network were analyzed by CytoScape. After marking gastric cancer-related genes extracted from the CancerResource, GeneRIF, and COSMIC databases, the FANMOD software was used for the mining of gastric cancer-related motifs in a network with three vertices. The significant motif difference method was adopted to identify significantly different motifs in the normal and cancer states. Finally, we conducted a series of analyses of the significantly different motifs, including gene ontology, function annotation of genes, and model classification. A human signaling network was constructed, with 1643 nodes and 5089 regulating interactions. The network was configured to have the characteristics of other biological networks. There were 57,942 motifs marked with gastric cancer-related genes out of a total of 69,492 motifs, and 264 motifs were selected as significantly different motifs by calculating the significant motif difference (SMD) scores. Genes in significantly different motifs were mainly enriched in functions associated with cancer genesis, such as regulation of cell death, amino acid phosphorylation of proteins, and intracellular signaling cascades. The top five significantly different motifs were mainly cascade and positive feedback types. Almost all genes in the five motifs were cancer related, including EPOR, MAPK14, BCL2L1, KRT18, PTPN6, CASP3, TGFBR2, AR, and CASP7. The development of cancer might be curbed by inhibiting signal transductions upstream and downstream of the selected motifs.


Assuntos
Humanos , Mineração de Dados , Redes Reguladoras de Genes , Transdução de Sinais/genética , Neoplasias Gástricas/genética , Motivos de Aminoácidos/genética , Morte Celular , Carcinogênese/genética , Retroalimentação Fisiológica , Regulação da Expressão Gênica , Ontologia Genética , Anotação de Sequência Molecular , Fosforilação , Neoplasias Gástricas/metabolismo
4.
Braz. j. med. biol. res ; 47(10): 834-841, 10/2014. tab, graf
Artigo em Inglês | LILACS | ID: lil-722173

RESUMO

In this study, biomarkers and transcriptional factor motifs were identified in order to investigate the etiology and phenotypic severity of Down syndrome. GSE 1281, GSE 1611, and GSE 5390 were downloaded from the gene expression ominibus (GEO). A robust multiarray analysis (RMA) algorithm was applied to detect differentially expressed genes (DEGs). In order to screen for biological pathways and to interrogate the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database, the database for annotation, visualization, and integrated discovery (DAVID) was used to carry out a gene ontology (GO) function enrichment for DEGs. Finally, a transcriptional regulatory network was constructed, and a hypergeometric distribution test was applied to select for significantly enriched transcriptional factor motifs. CBR1, DYRK1A, HMGN1, ITSN1, RCAN1, SON, TMEM50B, and TTC3 were each up-regulated two-fold in Down syndrome samples compared to normal samples; of these, SON and TTC3 were newly reported. CBR1, DYRK1A, HMGN1, ITSN1, RCAN1, SON, TMEM50B, and TTC3 were located on human chromosome 21 (mouse chromosome 16). The DEGs were significantly enriched in macromolecular complex subunit organization and focal adhesion pathways. Eleven significantly enriched transcription factor motifs (PAX5, EGR1, XBP1, SREBP1, OLF1, MZF1, NFY, NFKAPPAB, MYCMAX, NFE2, and RP58) were identified. The DEGs and transcription factor motifs identified in our study provide biomarkers for the understanding of Down syndrome pathogenesis and progression.


Assuntos
Animais , Humanos , Camundongos , Ratos , Motivos de Aminoácidos/genética , Biologia Computacional/métodos , Síndrome de Down/genética , Redes Reguladoras de Genes/genética , Fatores de Transcrição/análise , Algoritmos , Biomarcadores/análise , Bases de Dados Genéticas , Síndrome de Down/etiologia , Expressão Gênica , Ontologia Genética , Anotação de Sequência Molecular/métodos , Fenótipo , Análise Serial de Proteínas/métodos , Regulação para Cima/genética
5.
Braz. j. infect. dis ; 16(3): 250-255, May-June 2012. tab
Artigo em Inglês | LILACS | ID: lil-638558

RESUMO

OBJECTIVE: This study aimed to determine the natural prevalence of variants of tyrosine-methionine-aspartic acid-aspartic acid (YMDD) motif in patients with chronic hepatitis B (CHB), and to explore its relation with demographic and clinical features, hepatitis B virus (HBV) genotypes, and HBV DNA levels. METHODS: A total of 1,042 antiviral treatment naïve CHB patients (including with lamivudine [LAM]) in the past year were recruited from outpatient and inpatient departments of six centers from December 2008 to June 2010. YMDD variants were analyzed using the HBV drug resistance line probe assay (Inno-Lipa HBV-DR). HBV genotypes were detected with polymerase chain reaction (PCR) microcosmic nucleic acid cross-ELISA, and HBV deoxyribonucleic acid (DNA) was quantitated with real-time PCR. All serum samples underwent tests for HBV, HCV, and HDV with ELISA. RESULTS: YMDD variants were detected in 23.3% (243/1042) of CHB patients. YMDD mutation was accompanied by L180M mutation in 154 (76.9%) patients. Both wild-type HBV and YMDD variant HBV were present in 231 of 243 patients. Interestingly, 12 patients had only YIDD and/or YVDD variants without wild YMDD motif. In addition, 27.2% (98/359) of HbeAg-positive patients had YMDD mutations, which was higher than that in HbeAg-negative patients (21.2%, 145/683). The incidence of YMDD varied among patients with different HBV genotypes, but the difference was not significant. Moreover, the incidence of YMDD in patients with high HBV DNA level was significantly higher than that in those with low HBV DNA level. CONCLUSION: Mutation of YMDD motif was detectable at a high rate in CHB patients in this study. The incidence of YMDD may be correlated with HBeAg and HBV DNA level.


Assuntos
Adulto , Feminino , Humanos , Masculino , Antivirais/uso terapêutico , Ácido Aspártico/genética , Vírus da Hepatite B/genética , Hepatite B Crônica/tratamento farmacológico , Metionina/genética , Mutação/genética , Tirosina/genética , Motivos de Aminoácidos/efeitos dos fármacos , Motivos de Aminoácidos/genética , DNA Viral/análise , Genótipo , Vírus da Hepatite B/efeitos dos fármacos , Hepatite B Crônica/virologia , Reação em Cadeia da Polimerase
7.
Braz. j. med. biol. res ; 40(12): 1605-1614, Dec. 2007. graf, tab
Artigo em Inglês | LILACS | ID: lil-466741

RESUMO

Given the loss of therapeutic efficacy associated with the development of resistance to lamivudine (LMV) and the availability of new alternative treatments for chronic hepatitis B patients, early detection of viral genotypic resistance could allow the clinician to consider therapy modification before viral breakthrough and biochemical relapse occur. To this end, 28 LMV-treated patients (44 ± 12 years; 24 men), on their first therapy schedule, were monitored monthly at four Brazilian centers for the emergence of drug resistance using the reverse hybridization-based INNO-LiPA HBV DR assay and occasionally sequencing (two cases). Positive viral responses (HBV DNA clearance) after 6, 12, and 18 months of therapy were achieved by 57, 68, and 53 percent of patients, while biochemical responses (serum alanine aminotransferase normalization) were observed in 82, 82, and 53 percent of cases. All viral breakthrough cases (N = 8) were related to the emergence of YMDD variants observed in 7, 21, and 35 percent of patients at 6, 12, and 18 months, respectively. The emergence of these variants was not associated with viral genotype, HBeAg expression status, or pretreatment serum alanine aminotransferase levels. The detection of resistance-associated mutations was observed before the corresponding biochemical flare (41 ± 14 and 60 ± 15 weeks) in the same individuals. Then, if highly sensitive LMV drug resistance testing is carried out at frequent and regular intervals, the relatively long period (19 ± 2 weeks) between the emergence of viral resistance and the onset of biochemical relapse can provide clinicians with ample time to re-evaluate drug therapy.


Assuntos
Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Motivos de Aminoácidos/genética , Farmacorresistência Viral/genética , Vírus da Hepatite B/genética , Hepatite B Crônica/tratamento farmacológico , Lamivudina/uso terapêutico , Inibidores da Transcriptase Reversa/uso terapêutico , Alanina Transaminase/sangue , DNA Viral/sangue , Seguimentos , Antígenos E da Hepatite B/sangue , Vírus da Hepatite B/efeitos dos fármacos , Vírus da Hepatite B/imunologia , Hepatite B Crônica/imunologia , Hepatite B Crônica/virologia , Mutação/genética , Reação em Cadeia da Polimerase , Estudos Prospectivos
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